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Plant resistance against insects mainly depends on nutrient restriction and toxic metabolites, but the relative importance of nutrition and toxins remains elusive. We examined performance, nutrition ingestion, and transcriptome response of the English grain aphid, Sitobion avenae, feeding on resistant Xiaoyan 22 (XY22) and susceptible Xinong 979 (XN979) wheat cultivars. Aphids had lower body weight and fecundity when feeding on XY22 than on XN979, although the phloem sap of XY22 had a higher nutritive quality (in terms of amino acid:sucrose ratio). Aphids feeding on XY22 also had a lower honeydew excretion rate than those on XN979, suggesting that aphids ingested less phloem sap from XY22. The transcriptome data showed 600 differentially expressed genes (DEGs), and 11 of the top 20 KEGG pathways significantly enriched in DEGs were involved in nutrient metabolism. We found 81 DEGs associated with the metabolism of sugars, lipids, and amino acids, 59 of which were significantly downregulated in aphids feeding on XY22. In contrast, there were 18 DEGs related to detoxifying metabolism, namely eight UDP-glucuronosyltransferases, six cytochromes P450 monooxygenases, one glutathione S-transferase, two ATP-binding cassette transporters, and one major facilitator superfamily transporter; 12 of these were upregulated in the aphids feeding on XY22. Our results indicated that both the quantity and quality of phloem nutrition available to aphids are critical for the growth and development of aphids, and the higher resistance of XY22 is mainly due to the reduction in phloem sap ingested by aphids, rather than toxic metabolites.

Insect cuticular hydrocarbons (CHCs) serve as important intersexual signaling chemicals and generally show variation between the sexes, but little is known about the generation of sexually dimorphic hydrocarbons (SDHCs) in insects. In this study, we report the molecular mechanism and biological significance that underlie the generation of SDHC in the German cockroach Blattella germanica. Sexually mature females possess more C29 CHCs, especially the contact sex pheromone precursor 3,11-DimeC29. RNA interference (RNAi) screen against the fatty acid elongase family members combined with heterologous expression of the genes in yeast revealed that both BgElo12 and BgElo24 were involved in hydrocarbon (HC) production, but BgElo24 is of wide catalytic activities and is able to provide substrates for BgElo12, and only the female-enriched BgElo12 is responsible for sustaining female-specific HC profile. Repressing BgElo12 masculinized the female CHC profile, decreased contact sex pheromone level, and consequently reduced the sexual attractiveness of female cockroaches. Moreover, the asymmetric expression of BgElo12 between the sexes is modulated by sex differentiation cascade. Specifically, male-specific BgDsx represses the transcription of BgElo12 in males, while BgTra is able to remove this effect in females. Our study reveals a novel molecular mechanism responsible for the formation of SDHCs and also provide evidences on shaping of the SDHCs by sexual selection, as females use them to generate high levels of contact sex pheromone., Sexual dimorphism of body waxes is prevalent in insects; this study reveals that the sex-differentiation pathway regulates fatty acid elongation, ensuring production of the sexually dimorphic cuticular hydrocarbons needed for high levels of sex pheromone and sexual attractiveness in female cockroaches.

Water retention is critical for physiological homeostasis and survival in terrestrial insects. While deposition of hydrocarbons on insect cuticles as a key measure for water conservation has been extensively investigated, we know little about other mechanisms for preventing water loss in insects. Here, we report two fatty acid synthetic genes that are independent of hydrocarbon production but crucial for water retention in the German cockroach Blattella germanica (L.). First, an integument enriched fatty acid elongase gene (BgElo1) was identified as a critical gene for desiccation resistance in B. germanica; however, knockdown of BgElo1 surprisingly failed to cause a decline in cuticular lipids. In addition, RNA interference (RNAi)-knockdown of an upstream fatty acid synthase gene (BgFas3) showed a similar phenotype, and transmission electron microscopy analysis revealed that BgFas3- or BgElo1-RNAi did not affect cuticle architecture. Bodyweight loss test showed that repression of BgFas3 and BgElo1 significantly increased the weight loss rate, but the difference disappeared when the respiration was closed by freeze killing the cockroaches. A water immersion test was performed, and we found that BgFas3- and BgElo1-RNAi made it difficult for cockroaches to recover from drowning, which was supported by the upregulation of hypoxia-related genes after a 10-h recovery from drowning. Moreover, a dyeing assay with water-soluble Eosin Y showed that this was caused by the entry of water into the respiratory system. Our research suggests that BgFas3 and BgElo1 are required for both inward and outward waterproofing of the respiratory system. This study benefits the understanding of water retention mechanisms in insects.

Free amino acids in the phloem sap are the dominant nitrogen source for aphids, but their availability is usually poor. Although some studies have explored the effect of dietary amino acid restriction on aphid performance, little is known about the molecular basis of these effects. Here, we examined the performance and transcriptome of the green peach aphid, Myzus persicae, fed a standard diet (Control diet) or a diet containing 50% of the total amino acids of the Control diet (Half diet). Aphid weight and fecundity were significantly reduced in the Half diet group. Transcriptomic analysis showed that a total of 1460 genes were differentially expressed between the groups were fed on the two diets, which many of them were associated with nutrient and energy metabolism. When feeding on the Half diet, aphids upregulated genes associated with the amino acid biosynthetic pathway (predominantly amino acid biosynthesis genes and some amino acid transporter genes) as well as the cysteine and serine protease genes. Furthermore, these aphids displayed increased expression of genes associated with glycolysis, which could generate intermediates for de novo amino acid biosynthesis. Consistent with this, elevated glucose levels were observed in aphids in the Half diet group. Additionally, the expression levels of several genes associated with hormonal signaling pathway were altered. Several genes related to juvenile hormone and insulin-like peptide (ILP) signaling were downregulated, including Krüppel homolog 1 (Kr-h1) and insulin-like peptide 5 (Ilp5), respectively. In contrast, several genes related to ecdysone signaling were upregulated including broad-complex core protein (Br-c) and shade (Shd). Despite their poor performances, M. persicae adapted to dietary restriction of amino acids, through upregulation of genes involved in amino acid biosynthesis, glycolysis, and protein degradation, as well as by altering the expression level of genes involved in hormone signaling pathways.

Armyworm feeding in large, destructive groups is hugely difficult to control and the oriental armyworm, Mythimna separata (Walk), is one such pest. In this study, we reported a semisynthetic artificial diet for the oriental armyworm. This diet is based on Ritter’s diet, a formula developed for Heliothis zea. The survival of M. separata was extremely low and only around 2% insects can reach the adult stage on Ritter’s diet. But, it can reach up to 100% if corn leaf powder (CLP) was mixed, and insects grew faster and gained more mass. After testing a set of mixtures of Ritter’s diet and CLP, we found that 14.3% was the optimal proportion of CLP for making the artificial diet. We then used chloroform to extract CLP. Insect performance was still much better on Ch-extracted CLP diets than that on Ritter’s diet, but it was poorer than that on the diets containing unprocessed CLP, suggesting that the essential factor(s) was only partially extracted from corn leaf. We then used methanol and dichloromethane, two solvents differing in their polarity, to process the extractions and analyzed the extracted chemicals using gas chromatography–mass spectrometry (GC-MS). Insects had a better performance on dichloromethane-extracted CLP diet in comparison to methanol-extracted one, indicating that the important factor(s) is more prone to methanol extraction. The reported recipe here is useful for the research on M. separata and possibly other grain-crop eating armyworms. The functions of the chemicals extracted from corn leaf tissue can be investigated in the future studies.

Plants emit volatile organic compounds (VOCs) in response to herbivore attack. VOCs emitted from the Chinese cabbage cultivars in response to the damage by the green peach aphid, Myzus persicae, were unknown. Using a solid-phase microextraction-based headspace collection method, we investigated and compared the emissions of VOCs from seven Chinese cabbage cultivars (Qibao, Qingan 80, Lvlong, Yuanbao, Qingan 70, Jinlv, and Lvqiu 66) in response to M. persicae attack. Our results showed that the VOCs emitted from the cultivars Qingan 80 and Yuanbao differed significantly from the other cultivars in response to the attraction of wingless M. persicae. Most importantly, out of the 27 detected compounds, α-caryophyllene was detected only in Qingan 80 and Qibao, but not in the other five cultivars. Among the compounds detected, 2 monoterpene and 12 terpenes were predominant in all cabbage cultivars. Furthermore, the wingless M. persicae showed preference to Qingan 80 while it had the highest nitrogen content among the tested cultivars. Moreover, we found a remarkable relationship among M. persicae attraction, plant nitrogen content, and total volatile emissions. Nitrogen content of the plants has a significant impact on volatile emission and preference behavior of M. persicae. Our results indicate that the wingless M. persicae were efficient in their interspecific host selection with an ability to distinguish plant cultivar differences by leaf nitrogen content. This study will be helpful in understanding aphid host selection, and sets a stage to further study the attractant-based integrated aphid management programs.

Rising atmospheric carbon dioxide concentration ([CO 2 ]) and warming are two of the most conspicuous characteristics of global climate change in this century, and they are responsible for a variety of physiological responses of plants. However, studies addressing the combined impact of [CO 2 ] and temperature on plant physiological responses are still scarce. We studied the combined effects of long term elevated [CO 2 ] and temperature on dynamic changes of physiological metabolism in cotton with growth and development. Dry matter content (DMC), N concentration, C/N ratio, soluble sugar content, starch content, total nonstructural carbohydrate (TNC), total phenolic concentration and amino acid levels in cotton leaves in response to long term [CO 2 ] and temperature levels were assayed during different growth times. Elevated [CO 2 ] and temperature independently increased DMC at different growth times but the highest increase of DMC occurred under both elevated [CO 2 ] and elevated temperature. Elevated [CO 2 ] reduced leaf N concentration and increased C/N ratio; however, no differences were found at ambient or elevated temperature after both 75 and 105 days of treatments. The leaves grown in elevated [CO 2 ] had higher soluble sugar, starch, TNC contents and total phenolic concentration, whereas lower levels of these components were observed at a higher temperature. There were different interactive effects between [CO 2 ] and temperature on these physiological characteristics at different growth times. Elevated [CO 2 ] increased amino acids of Asp, His, Ile and Thr levels and reduced Glu, Ala and Pro levels at different growth times under ambient temperature. Under elevated temperature, [CO 2 ] enrichment increased Glu and Thr levels, whereas it reduced Ala, Asp and Val levels. There was a significant interactive effect between [CO 2 ] and temperature on 8, 9 and 11 out of the 15 amino acids after 45, 75 and 105 days of treatment, respectively. The results indicate these physiological characteristics in the cotton leaves during different growth times will change differently in response to individual environmental factor or the combination of different environmental factors, and the physiological metabolism in cotton under long term elevated [CO 2 ] and temperature will dynamically change with growth and development, thus they may have different metabolic mechanisms to deal with future climate change. Further work is required to elucidate the mechanisms underlying an interactive effect of [CO 2 ] and temperature on plant physiological and metabolic change as well as the effect of plant nutritional quality changes on plant-herbivore interaction.

Melanism in insects is important for their physical protection, immunoreactions, and sclerotization. The vetch aphid, Megoura viciae (Buckton), has relatively strong tanning in its prothorax, head, antennae, cornicles, and legs. It was hypothesized that M. viciae may sequester the high level of l -DOPA in its host Vicia faba to help in its melanization process for ecological adaptation. To confirm this hypothesis, the amount of l -DOPA in M. viciae was modified and quantified. We first generated a Trifolium repens (clover, low l -DOPA containing) host to cut off the extra l -DOPA intake by M. viciae. The rate-limiting tyrosine hydroxylase gene of M. viciae (MV-TH) was then cloned and analyzed. To further reduce the l -DOPA level in the insect, RNAi was used to downregulate the transcriptional level of MV-TH. Our results confirmed that M. viciae could indeed sequester l -DOPA in its body, and its ample storage of this amino acid could be the reason for the strong tanning of its body. M. viciae reared on T. repens could upregulate its MV-TH to enhance l -DOPA biosynthesis and thus maintain a high level of l -DOPA. The MV-TH repression by RNAi lasted for about 3 days, successfully decreasing the l -DOPA level. Aside from a slight decrease in exuvia tanning, no other obvious change in body appearance was detected in the RNAi-treated insect. Although M. viciae can obtain most of its l -DOPA directly from its original host, its internal l -DOPA synthetase is still functional, especially when extra l -DOPA is removed from the diet. This capability to enhance its shield ensures the ecological adaptation of this insect.

In natural ecological systems, plants are often simultaneously attacked by both insects and pathogens, which can affect each other's performance and the interactions can be extended to higher trophic levels, such as parasitoids. The English grain aphid (Sitobion avenae) and powdery mildew (Blumeria graminis f. sp. tritici) are two common antagonists that pose a serious threat to wheat production. Numerous studies have investigated the effect of a single factor (insect or pathogen) on wheat production. However, investigation on the interactions among insect pests, pathogens, and parasitoids within the wheat crop system are rare. Furthermore, the influence of the fungicide, propiconazole, has been found to imitate the natural ecosystem. Therefore, this study investigated the effects of B. graminis on the biological performance of grain aphids and the orientation behavior of its endoparasitic wasp Aphidius gifuensis in the wheat system. Our findings indicated that B. graminis infection suppressed the feeding behavior, adult and nymph weight, and fecundity and prolonged the developmental time of S. avenae. We found that wheat host plants had decreased proportions of essential amino acids and higher content of sucrose following aggravated B. graminis infection. The contents of Pro and Gln increased in the wheat plant tissues after B. graminis infection. In addition, B. graminis infection elicited immune responses in wheat: increase in the expression of defense genes, content of total phenolic compounds, and activity of three related antioxidant enzymes. Moreover, co-infection of B. graminis and S. avenae increased the attraction to A. gifuensis compare to that after infestation with aphids alone. In conclusion, our results indicated that B. graminis infection adversely affected the performance of S. avenae in wheat through restricted nutrition and induced defense response. Furthermore, the preference of parasitoids in such an interactive environment might provide an important basis for pest management control.

The maternal effects of the English grain aphid, Sitobion avenae on offspring phenotypes and performance on wheat varieties with different resistance traits were examined. We found that both conditioning wheat varieties(the host plant for over 3 months) and transition wheat varieties affected the biological parameters of aphid offspring after they were transferred between wheat varieties with different resistance traits. The conditioning varieties affected weight gain, development time (DT), and the intrinsic rate of natural increase (rm), whereas transition varieties affected the fecundity, rm, net reproductive rate, and fitness index. The conditioning and transition wheat varieties had significant interaction effects on the aphid offspring’s DT, mean relative growth rate, and fecundity. Our results showed that there was obvious maternal effects on offspring when S. avenae transferred bwteen wheat varieties with different resistance level, and the resistance traits of wheat varieties could induce an interaction between the conditioning and transition wheat varieties to influence the growth, development, reproduction, and even population dynamics of S. avenae. The conditioning varieties affected life-history traits related to individual growth and development to a greater extent, whereas transition varieties affected fecundity and population parameters more.

Plant leaves of different ages differ in nutrients and toxic metabolites and thus exhibit various resistance levels against insect herbivores. However, little is known about the influence of leaf ontogeny on plant resistance to phloem-feeding insects. In this study, we found that the green peach aphid, Myzus persicae, preferred to settle on young cabbage leaves compared with mature or old leaves, although young leaves contained the highest concentration of glucosinolates. Furthermore, aphids feeding on young leaves had higher levels of glucosinolates in their body, but aphids performed better on young leaves in terms of body weight and population growth. Phloem sap of young leaves had higher amino acid:sugar molar ratio than mature leaves, and aphids feeding on young leaves showed two times longer phloem feeding time and five times more honeydew excretion than on other leaves. These results indicate that aphids acquired the highest amount of nutrients and defensive metabolites when feeding on young cabbage leaves that are strong natural plant sinks. Accordingly, we propose that aphids generally prefer to obtain more nutrition rather than avoiding host plant defense, and total amount of nutrition that aphids could obtain is significantly influenced by leaf ontogeny or source-sink status of feeding sites.

The behaviour of the greenhouse whitefly, Trialeurodes vaporariorum Westwood (Hemiptera: Aleyrodidae), is known to be affected by plant volatile cues, but its attraction or repellent to specific volatile cues has not been deeply studied yet. Therefore, the aim of our study was to identify the most attractive plant among cultivars of tomato (Solanum lycopersicum) and eggplant (Solanum melongena) to evaluate the volatiles of plants to identify the chemical compound(s) that attract T. vaporariorum. We speculated that whitefly–host plant interaction primarily depends on plant volatile emissions and that once the plant is damaged, it might attract more whiteflies. Three intact (uninfested) tomato, four intact eggplant cultivars and whitefly-infested plants of the most whitefly attractive tomato and eggplant cultivars were examined by behavioral assay experiments for attractiveness to T. vaporariorum and headspace volatile were determined by Solid-phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS). Whiteflies had the highest preference for the intact eggplant Kuai Yuan Qie (KYQ) among the eggplant and the tomato plant cultivars in bioassay experiments. Although both male and female whiteflies were significantly more attracted to infested KYQ plants than to intact plants, whitefly females did not select conspecific-infested YG plants. The volatile emissions among different plant cultivars in individual species and infested versus intact plants were significantly different. Among these volatiles, identified major green leaf volatile (GLVs) ((Z)-3-hexen-1-ol) and terpenoids (α-pinene, (E)-β-caryophyllene, α-humulene, azulene) showed a constitutive relationship with the most whitefly preference plants. Our findings provide new insights into the chemical compounds that attract or repel whiteflies.

Tomato (Solanum lycopersicum) production is seriously hampered by the infestation of the sweetpotato whitefly, Bemisia tabaci MEAM 1 (Middle East-Asia Minor 1). The infestation behavior of the whiteflies could be affected by the quantity of plant released volatile organic compounds (VOCs) related to nitrogen concentrations of the plant. In this study, we determined the infestation behavior of B. tabaci to the tomato plants that produced different levels of VOCs after application of different levels of nitrogen with a wind tunnel and an olfactometer. We also analyzed the VOCs released from nitrogen-treated tomato plants using solid phase microextraction and gas chromatography-mass spectrometry. The results revealed that the production of eight VOCs (β-pinene, (+)-4-carene, α-terpinene, p-cymene, β-phellandrene, α-copaene, β-caryophyllene, and α-humulene) was reduced after the plants were treated with high levels of nitrogen. However, more whiteflies were attracted to the tomato plants treated with high levels of nitrogen than to the plants treated with normal or below normal levels of nitrogen. These results clearly indicated that nitrogen can change the quality and quantity of tomato plant volatile chemicals, which play important roles in B. tabaci host plant selection.

The green peach aphid, Myzus persicae Sulzer, is a notorious pest on vegetables, which often aggregates in high densities on crop leaves. In this study, we investigated whether M. persicae could suppress the resistance level of Chinese cabbage Brassica pekinensis. M. persicae performed better in terms of weight gain (~33% increase) and population growth (~110% increase) when feeding on previously infested (pre-infested) Chinese cabbage compared with those on non-infested plants. However, when given a choice, 64% of the aphids preferred to settle on non-infested leaves, while 29% of aphids chose pre-infested leaves that had a 2.9 times higher concentration of glucosinolates. Aphid feeding significantly enhanced the amino acid:sugar ratio of phloem sap and the absolute amino acid concentration in plant leaves. Aphid infestation significantly increased the expression levels of salicylic acid (SA) marker genes, while it had marginal effects on the expression of jasmonate marker genes. Exogenously applied SA or methyl jasmonate had no significant effects on M. persicae performance, although these chemicals increased glucosinolates concentration in plant leaves. M. persicae infestation increase amino acid:sugar ratio and activate plant defenses, but aphid performed better on pre-infested plants, suggesting that both nutrition and toxics should be considered in insect-plant interaction.

Wingless forms of aphids are relatively sedentary, and have a limited ability to migrate or disperse. However, they can drop off hosts or walk away if disturbed, or their food quality or quantity become deteriorated. Earlier, we found that the pea aphid, Acyrthosiphon pisum (Harris, 1776), could use differed strategies to escape danger and locate new host plants. To determine the mechanisms behind the different strategies, we undertook a series of studies including the aphids' host location, energy reserves under starvation, glycogenesis, sugar assimilation, olfactory and probing behaviors. We found that in our controlled laboratory conditions, one strain (local laboratory strain) moved longer distances and dispersed wider ranges, and correspondingly these aphids assimilated more sugars, synthesized more glycogen, and moved faster than another strain (collected from Gansu Province, northwestern China). However, the latter strain could locate the host faster, probed leaves more frequently, and identified plant leaves more accurately than the former strain after they were starved. Our results explained how flightless or wingless insects adapt to fit biotic and abiotic challenges in the complex processes of natural selection., Summary: Two strains of pea aphid (Acyrthosiphon pisum) show different dispersal strategies, glycogen level and probing behaviors. One strain [Yangling strain (YL)] moves longer and disperses wider than another strain [Lanzhou (LZ)] based on higher glycogen. LZ strain probed more frequently, and identified plant more accurately than YL strain.

Terrestrial insects deposit a layer of hydrocarbons (HCs) as waterproofing agents on their epicuticle. The insect-specific CYP4G genes, subfamily members of P450, have been found in all insects with sequenced genomes to date. They are critical for HC biosynthesis in Drosophila; however, their functional roles in other insects including the piercing-sucking hemipterous aphids remain unknown. In this study, we presented the molecular characterization and a functional study of the CYP4G51 gene in the pea aphid, Acyrthosiphon pisum (Harris). CYP4G51 transcript was detectable across the whole life cycle of A. pisum, and was prominently expressed in the aphid head and abdominal cuticle. Up-regulation of CYP4G51 under desiccation stress was more significant in the third instar nymphs compared with the adults. Also, up-regulation of CYP4G51 was observed when the aphids fed on an artificial diet compared with those fed on the broad bean plant, and was positively correlated with a high level of cuticular HCs (CHCs). RNAi knockdown of CYP4G51 significantly reduced its expression and caused reductions in both internal and external HCs. A deficiency in CHCs resulted in aphids being more susceptible to desiccation, with increased mortality under desiccation stress. The current results confirm that CYP4G51 modulates HC biosynthesis to protect aphids from desiccation. Moreover, our data also indicate that saturated and straight-chain HCs play a major role in cuticular waterproofing in the pea aphid. A. pisum CYP4G51 could be considered as a novel RNAi target in the field of insect pest management.

Herbivores can ingest and store plant-synthesized toxic compounds in their bodies and sequester those compounds for their own benefits. The broad bean, Vicia faba L., contains a high quantity of L-DOPA (L-3,4-dihydroxyphenylalanine), which is toxic to many insects. However, the pea aphid, Acyrthosiphon pisum, can feed on V. faba normally, whereas many other aphid species could not. In this study, we investigated how A. pisum utilizes plant-derived L-DOPA for their own benefit. L-DOPA concentrations in V. faba and A. pisum were analyzed to prove L-DOPA sequestration. L-DOPA toxicity was bioassayed using an artificial diet containing high concentrations of L-DOPA. We found that A. pisum could effectively adapt and store L-DOPA, transmit it from one generation to the next. We also found that L-DOPA sequestration verity differed in different morphs of A. pisum. After analyzing the melanization efficiency in wounds, mortality and deformity of the aphids at different concentrations of L-DOPA under ultraviolet radiation (UVA 365.0 nm for 30 min), we found that A. pisum could enhance L-DOPA assimilation for wound healing and UVA-radiation protection. Therefore, we conclude that A. pisum could acquire L-DOPA and use it to prevent UVA damage. This study reveals a successful co-evolution between A. pisum and V. faba.

The green peach aphid, Myzus persicae Sulzer, is a notorious pest on vegetables, which often aggregates in high densities on crop leaves. In this study, we investigated whether M. persicae could suppress the resistance level of Chinese cabbage Brassica pekinensis. M. persicae performed better in terms of weight gain (~33% increase) and population growth (~110% increase) when feeding on previously infested (pre-infested) Chinese cabbage compared with those on non-infested plants. However, when given a choice, 64% of the aphids preferred to settle on non-infested leaves, while 29% of aphids chose pre-infested leaves that had a 2.9 times higher concentration of glucosinolates. Aphid feeding significantly enhanced the amino acid:sugar ratio of phloem sap and the absolute amino acid concentration in plant leaves. Aphid infestation significantly increased the expression levels of salicylic acid (SA) marker genes, while it had marginal effects on the expression of jasmonate marker genes. Exogenously applied SA or methyl jasmonate had no significant effects on M. persicae performance, although these chemicals increased glucosinolates concentration in plant leaves. M. persicae infestation increase amino acid:sugar ratio and activate plant defenses, but aphid performed better on pre-infested plants, suggesting that both nutrition and toxics should be considered in insect-plant interaction.